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Poly(A) Polymerase Tailing Kit

Poly(A) Polymerase Tailing Kit

Catalog Number: UA070116 Brand: UA BIOSCIENCE
Price:
Regular price $420 USD
Regular price Sale price $420 USD
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Product Details

Product Specification


Stability & Storage

Store at -25 ~ -15℃ for 2 years

Reference


Components

组分

UA070116-50 Rxns

UA070116-250 Rxns

E. coli Poly(A) Polymerase

 (5 U/µl)

100 μl

 5 × 100 μl

10 X PAP Reaction Buffer

300 μl

5 × 300 μl

10 mM ATP

300 μl

5 × 300 μl

RNase Free dH2O

1 ml

5 × 1 ml


Protocol

1. Prepare the following reaction system on ice according to the table below:

Component

Volume

Final Concentration

10 X PAP Reaction Buffer

5 μl

1 X

RNase Inhibitor (40U/μl)

1.25 μl

1 U/μl (Optional)

E.coli Poly(A) Polymerase

(5 U/μl)

2 μl

0.2 U/μl

10 mM ATP

5 μl

1 mM

RNase Free dH2O

Up to 50 μl

-

Gently pipette to mix thoroughly. 2. Poly(A) tailing reaction: Incubate at 37°C for 30–60 min. 3. Reaction termination: Immediately store at –20°C after completion, or add EDTA to a final concentration of 10 mM, or perform phenol/chloroform extraction followed by ammonium salt/ethanol precipitation of RNA.

Guidelines

1. Due to the involvement of RNA manipulation, strict adherence to RNA operation protocols is required to avoid RNase contamination. Related reagents and consumables must be treated with DEPC to remove RNase or ensured to be RNase-free. 2. The RNA used for the tailing reaction should be appropriately purified before use and dissolved in RNase-free dH2O. The solution should not contain EDTA or salts. 3. If it is difficult to ensure a strictly RNase-free environment, it is recommended to add an appropriate amount of RNase Inhibitor to the reaction system to enhance the stability of RNA in the solution. 4. During experimental operations, enzyme products should be kept on ice at all times and stored at -20°C immediately after the experiment.

Picture

Bioactivity

Poly(A) Polymerase Tailing Kit对单链RNA添加Poly(A)尾的效果图。在50μl反应体系中,加入5 μg 541nt的单链RNA,以及图中指定量的Poly(A) Polymerase,37ºC孵育30 min,65ºC孵育20min终止反应。以下为琼脂糖凝胶电泳检测效果图。
Lane 1阴性对照(未加Poly(A) Polymerase);
Lane 2 0.008 U PAP;
Lane 3 0.04 U PAP;
Lane 4 0.2 U PAP;
Lane 5 1 U PAP;
Lane 6 5 U PAP;
Lane 7 10 U PAP;